Cromatest HBsAg Test Device

PRINCIPLE

The LINEAR HBsAg cassette is  a lateral  flow chromatographic immunoassay for the qualitative detection of Hepatitis B surface antigen (HBsAg) in human serum or plasma at the level equal to or higher than 1 ng/ml. It is intended to be used as a screening test and as an aid in the diagnosis of infection with Hepatitis B virus (HBV). Any reactive specimen with the HBsAg cassette must be confirmed with alternative testing method(s) and clinical findings.

The LINEAR HBsAg cassette detects HBsAg through visual interpretation of color development on the internal strip. Anti- HBsAg antibodies are immobilized on the test region of the membrane. During testing, the specimen reacts with anti-HBsAg antibodies conjugated to colored particles and pre-coated onto the sample pad of the test. The mixture then migrates through the membrane by capillary action, and interacts with reagents on the membrane. If there is enough HBsAg in the specimen, a colored band will form at the test region of the membrane. The presence of this colored band indicates a positive result, while its absence indicates a negative result. The appearance of a colored band at the control region serves as a procedural control, indicating that the proper volume of specimen has been added and membrane wicking has occurred.

REAGENT COMPOSITION

HBsAg test device, contains protein A and anti-HBsAg antigen coated on the membrane.

STORAGE AND STABILITY

Store at 2-30ºC. The test cassette is stable through the expiration date printed on the sealed pouch. The test cassette must remain in the sealed pouch until use. DO NOT FREEZE. Do not use beyond the expiration date or devices with damaged pouch. Care should be taken to protect the components of the kit from contamination.

Discard If appear signs of deterioration:

• RPR-Carbon: Visible

Controls: Presence of particles and turbidity

SPECIMEN COLLECTION AND PREPARATION

• LINEAR HBsAg cassette is intended for use with human serum or plasma specimens only.
• Only clear, non-hemolyzed specimens are recommended for use with this test. Serum or plasma should be separated as soon as possible to avoid hemolysis.
• Perform testing immediately after specimen collection. Do not leave specimens at room temperature for prolonged periods. Specimens may be stored at 2-8°C for up to 3 days. For long term storage, specimens should be kept below -20°C.
• Bring specimens to room temperature prior to testing. Frozen specimens must be completely thawed and mixed well prior to Avoid repeated freezing and thawing of specimens.
• If specimens are to be shipped, pack them in compliance with all applicable regulations for transportation.
• Icteric, lipemic, hemolyzed, heat treated and contaminated sera may cause erroneous results.

MATERIAL REQUIRED

• Specimen collection
• General laboratory
• Timer

PROCEDURE

Allow test device, serum or plasma specimen, and/or controls to equilibrate to room temperature (15-30°C) prior to testing.

1. When ready to test, open the pouch at the notch and remove the Place the test device on a clean, flat surface.
2. Be sure to label the device with specimen’s ID
3. Fill the plastic dropper with the specimen. Holding the plastic dropper vertically, dispense 2 drops (about 60-90 µL) of specimen into the sample well (S). (Note)

Avoid trapping air bubbles in the specimen well (S), and do not add any solution to the result window.

4. As the test begins to work, color will migrate across the
5. Wait for the colored band(s) to appear. The result should be read at 15 minutes.

Do not read result after 20 minutes. To avoid confusion, discard the test device after interpreting the result.

INTERPRETATION OF RESULTS

POSITIVE: * Two distinct red lines appear. Two colored bands appear on the membrane. One band appears in the control region (C) and another band appears in the test region (T).

NEGATIVE: Only one colored band appears, in the control region (C). No apparent colored band appears in the test region (T).

INVALID: Control line fails to appear. Results from any test which has not produced a control band at the specified read time must be discarded. Please review the procedure and repeat with a new test. If the problem persists, discontinue using the kit immediately and contact your local distributor

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  QUALITY CONTROL

  An internal procedural control is included in the test. A colored line appearing in the control region (C) is considered an internal positive procedural control. It confirms sufficient specimen volume and correct procedural technique.

  External controls are not supplied with this kit. It is recommended that positive and negative controls should be tested as a good laboratory practice to confirm the test procedure and to verify proper test performance. Handle the negative and positive controls in the same manner as patient specimens.

  CLINICAL SIGNIFICANCE5

  Hepatitis B virus (HBV) is the prototypic member of the hepadnaviruses. Hepatitis B surface antigen (HBsAg) is located in the lipid envelope of this small DNA virus. During the replicative phase of the virus, this surface antigen is produced in excess and is detectable in the blood of the infected. The incubation period of HBV is 6 weeks to 6 months

  ANALYTICAL PERFORMANCE

  1. Clinical A total of 560 samples from susceptible subjects were tested with the Linear HBsAg Cassette and with a commercial HBsAg ELISA kit with a test sensitivity of 0.5 ng/mL. Comparison for all subjects is shown in the following table.
     

     HBsAg ELISA	Positive	Negative	Total
     Positive	97	0	97
     Negative	0	463	463
     Total	97	463	560

  2. Cross-Reactivity. Cross-reactivity with specimens from other infectious diseases
     

     Dengue Positive Serum	10	Negative
     HAV Positive Serum	10	Negative
     HCV Positive Serum	10	Negative
     HIV Positive Serum	10	Negative
     Syphilis Positive Serum	10	Negative
     TB Positive Serum	10	Negative
     H. pylori Positive Serum	10	Negative
     ANA Positive Serum	6	Negative
     HAMA Positive Serum	4	Negative
     RF Positive Serum (≤2,500 IU/ml)	3	Negative

     3.Interference. Common substances (such as pain and fever medication and blood components) may affect the performance of the Linear HBsAg Cassette. This was studied by spiking these substances into three levels of HBsAg standard controls. The results are presented in the following table and demonstrate that at the concentrations tested, the substances studied do not affect the performance of the Linear HBsAg Cassette.

  3. 1. This package insert must be read completely before performing the Failure to follow the insert gives inaccurate test results.
     2. Do not use the components in any other type of test kit as a substitute for the components in this kit.
     3. Do not use hemolized blood specimens for
     4. Wear protective clothing and disposable gloves while handling the kit reagents and clinical Wash hands thoroughly after performing the test.
     5. Users of this test should follow the US CDC Universal Precautions for prevention of transmission of HIV, HBV and other blood-borne
     6. Do not smoke, drink or eat in areas where specimens or kit reagents are being handled.
     7. Dispose of all specimens and materials used to perform the test as bio-hazardous waste.
     8. Handle the negative and positive controls in the same manner as patient specimens.
     9. Do not perform the test in a room with strong air flow, i.e. electric fan or strong air-conditioning.
        

        Potential Interfering Substances Spiked	HB	sAg Reactiv	ity
        	Negative	Weak Positive	Medium Positive
        Control	–	+	++
        Bilirubin 20 mg/dL	–	+	++
        Creatinine 442 µmol/L	–	+	++
        Glucose 55 mmol/L	–	+	++
        Albumin 50 g/L	–	+	++
        Salicylic Acid 4.34 mmol/L	–	+	++
        Heparin 3,000 U/L	–	+	++
        EDTA 3.4 µmol/L	–	+	++
        Human IgG 1,000mg/dL	–	+	++
        Sodium citrate 3.8%	–	+	++

        PRECAUTIONS

     LIMITATIONS OF PROCEDURE

     • The Assay Procedure and the Interpretation of Assay Result sections must be followed Failure to follow the procedure may give inaccurate results.
     • LINEAR HBsAg is a screening test. This screening test should be used for the detection of HBsAg in serum or plasma Neither the quantitative value nor the rate of HbsAg concentration can be determined by this qualitative test.
     • A non-reactive test result does not preclude the possibility of exposure to or infection with HBV.
     • LINEAR HBsAg will only indicate the presence of HBsAg in the specimen and should not be used as the sole criteria for the diagnosis of Hepatitis B viral
     • If the test result is negative and clinical symptoms persist, additional testing using other clinical methods is A negative result does not at anytime rule out the presence of HBsAg in blood, as HBsAg may be present below the minimum detection level of the test.
     • As with all diagnostic tests, a confirmed diagnosis should only be made by a physician after all clinical and laboratory findings have been evaluated.
     • Some specimens containing unusually high titers of heterophile antibodies or rheumatoid factor may affect expected results.

     • NOTE

       Add 1 drop of Saline or Phosphate-Saline buffer (common buffers used in clinic not provided in the kit) into the sample well if flow migration is not observed within 30 seconds in the result window, which could occur with a highly viscous specimen.

       NOTES

       1. The HBsAg Rapid Test is limited to the qualitative detection of HBsAg in human serum or plasma. The intensity of the test line does not have a linear correlation with the HBsAg titer in the specimenREFERENCES
       1. Emanuel Rubin and John Farber. The liver and biliary system. Acute viral hepatitis P 721-729. Rubin E, Farber JL Pathology 2nd ed. 1994. J.B. Lippincott, Philadelphia
       2. Kaplan PM, Greenman RL, Gerin JL, Purcell RH, Robinson DNA polymerase associated with human hepatitis B antigen. J Virol. 1973 12(5):995-1005.
       3. Dane DS, Cameron CH, Briggs Virus-like particles in serum of patients with Australia-antigen-associated hepatitis. Lancet. 1970;1(7649):695-8.
       4. Magnius LO, Espmark A. A new antigen complex co-occurring with Australia Acta Pathol Microbiol Scand [B] Microbiol Immunol. 1972;80(2):335-7